3.4.4 Cloning

• Clones are groups of genetically identical organisms, derived from a single original parent cell
• A cloned cell is a cell that is genetically identical to the cell that it originated from
• Sexual reproduction produces a zygote when gametes fuse
• In a single birth, this zygote is not cloned and will itself reproduce sexually as an adult
  • Identical twins are clones of each other as they are formed from one zygote splitting into two parts, which each develops into an embryo
• Clones form naturally and artificially
• The simple gardening technique of taking plant cuttings relies on cloning
• Other organisms are manipulated to form multiple clones when grown commercially eg. large-scale growth of crop plants, to ensure a uniform crop and good crop yields
• This ensures that desirable characteristics appear in the phenotypes of every organism

• Many plant species and some animal species have natural methods of cloning
• Asexual reproduction is much less common in animals than in plants
• Some small animals reproduce asexually by parthenogenesis eg. aphids
• The other naturally occurring incidence of cloning in animals is identical twins

Natural cloning in plants

• Many methods of cloning do not require seeds as it is not sexual reproduction that is occurring, it is asexual reproduction

• A well as runners, plants can propagate asexually using tubers, rhizomes, bulbs, suckers, and offsets
• All modes of natural plant cloning contain modified stems that can generate meristem tissue
• Potato tubers are swollen modified roots that form eyes on their surface
  • Eyes can sprout new growth (called 'chitting')
  • The starch stored in the tuber fuels the early growth of the new plant

• Ginger forms rhizomes, a modified stem that grows horizontally underground
  • New growth stems from nodes in the rhizome, forming new stems and adventitious roots
  • The section used in cookery is the rhizome

• Onions and garlic form bulbs that can grow adventitious roots underground and leafy shoots above ground
• Suckers are growths that appear from the root systems of many trees and shrubs, which can provide meristematic tissue for vegetative propagation
  • Examples are poplars, cherries and plums

• Offsets are small, virtually complete daughter plants that have been asexually produced on the mother plant
  • Examples are tulips and lilies

An example of natural cloning in plants with runners that form adventitious roots

Identical twins

• An egg is fertilised by a sperm as in a singleton birth
• This forms a zygote
• The single zygote undergoes a few cell cycles (mitotic divisions) to become an embryo
  • This is why identical twins are referred to as monozygotic
• At the embryo stage, the embryo splits in two; the exact causes of this kind of split are not well understood
• The two embryos that form are identical (have exactly the same genotype) and develop in utero (i.e. in the uterus) together
• The result is the birth of identical offspring, always of the same gender, with identical phenotypes
• Because non-identical twins are formed from separate eggs and sperm, they are not clones

Identical twins are natural clones when a zygote splits into two parts and each part develops into a separate embryo

• The process of embryo twinning (sometimes called splitting or fragmentation) produces offspring that are clones of each other but not of their parents
• It has been a routine procedure carried out to boost yields of livestock and promote desirable characteristics since the 1980s
• The key step is the deliberate division of the embryo into two half embryos
• Both halves contain cells that are pluripotent
  • The embryo is split at around the eight-cell stage
• These are then inserted into a surrogate mother for gestation and birth
• The surrogate gives birth to identical twins
• In some cases, embryos are split into single identical cells, each of which can be implanted into a separate surrogate mother animal
• Although embryo twinning guarantees desirable characteristics in the offspring, it is not possible to predict how many offspring will be produced within a herd of livestock, something of vital importance to a farmer

Embryo twinning of cattle by splitting the embryo

Therapeutic cloning

• This is a technique designed to use cloned cells to replace dead or damaged cells that cause a loss of function in an individual
• Embryos are cloned as in reproductive cloning, but the embryos are removed and subdivided
• Each individual embryo cell is pluripotent and can be cultured and artificially differentiated into any type of specialised cell
• In theory, any specialised cell can be derived by this method
  • Crucially, specialised cells with the same genome as the sufferer can be cloned and replaced

• An example is replacing specialised brain tissue in sufferers of Parkinson's Disease
• At present, there is a lot of potential for therapeutic cloning but little clinical progress has been made

• Methods have been developed for cloning adult animals using differentiated cells
• It would be desirable to clone a differentiated cell because by then it would be easy to assess the organism's characteristics and whether any of its traits were desirable enough to clone
  • Pluripotent cells can develop into any specialised cell, but it is difficult to predict whether, once differentiated, they will display desirable characteristics
• Differentiated cells are more difficult to clone because certain genes have been permanently switched off (those genes will never be transcribed again) as the cell has developed its specialised role
• However, pioneering work on Xenopus (the African clawed frog) in the 1950s involved:
  • Removal of nuclei from Xenopus tadpoles' somatic cells
  • It was significant that the somatic cell was already fully differentiated eg. a skin cell
  • Insertion of these nuclei into enucleated Xenopus egg cells (eggs whose own nuclei had been destroyed by UV radiation treatment)
• This resulted in embryos that grew, divided and differentiated into fully-functioning live tadpoles, and ultimately, adult Xenopus frogs
• Whilst this work was successful, cloning mammals from differentiated cells proved much more difficult
• The work on Xenopus frogs laid the platform for the first mammal cloned by nuclear extraction
• It wasn't until the 1990s that the first large mammal, Dolly the sheep, was cloned successfully

Cloning of Xenopus frogs by nuclear transfer from a differentiated cell

Production of cloned embryos produced by somatic cell nuclear transfer

• This is the method made famous by Dolly the sheep, cloned in Edinburgh, UK in 1996
• Its full name is Somatic Cell Nuclear Transfer (SCNT)
• Dolly made headlines as being the first livestock animal to be created from a clone
• Three separate animals are required:
  • The animal being cloned (by donating a cell)
  • The female to donate an egg cell
  • The surrogate mother

• How the procedure is carried out:
  • The animal to be cloned donates a somatic (body) cell
    • In Dolly's case, this was an udder cell
  • The egg cell is extracted from the egg donor and enucleated (its nucleus is removed by suction and discarded)
  • The nucleus from the udder cell is injected into the enucleated egg cell
  • The hybrid zygote cell is now treated to encourage it to divide by mitosis
  • The embryo is implanted into the surrogate mother for gestation and birth

Reproductive cloning of animals by somatic cell nuclear transfer